Development of the mature zebrafish as an animal model of human disease and aging.

Abstract

Zebrafish are a valuable model for developmental biology. However, mature zebrafish biology has not been explored, although many attributes suggest it will also be an important model for study of aging. This dissertation characterizes basic physiologic features and defines mechanisms of stress response of mature zebrafish. The hematological and biochemical parameters of mature zebrafish, including leukocyte differentials, total erythrocyte counts, and serum biochemical analytes, were within range of reported values for other fish species. These results provide reference values to compare with mutated zebrafish. To further develop zebrafish as models for human disease and aging, I characterized the heat shock response of zebrafish. Heat shock proteins (Hsps) are involved in many physiological processes and are diminished with age in other species. Accordingly, mature zebrafish were exposed to heat stress and various tissues analyzed for Hsp expression. Hsp70 and Hsp47 were upregulated in a tissue-specific manner, while Hsp90alpha, Hsp90beta, and Hsf1a were expressed but not upregulated in response to heat stress. A comparison of young versus mature zebrafish revealed decreased basal levels of Hsp70 and increased levels of Hsf1a in mature fish, demonstrating that there are age differences in their heat shock response, which supports use of mature zebrafish for study of heat shock response and aging. To further define the mechanism of heat shock response in zebrafish, I explored if heat stress-induces Hsp70 expression through ERK activation, as has been reported for mammals. Heat stress induced both Hsp70 expression and ERK1/2 phosphorylation in zebrafish Pac2 cells. Although ERK inhibitors PD98059 and U0126 blocked ERK1/2 phosphorylation, they did not block heat stress-induced Hsp70 expression. These results demonstrate that heat stress induces Hsp70 expression independent of ERK activation in Pac2 cells. This observation suggests that the heat shock response in zebrafish utilizes a different signaling pathway from that of mammals. In conclusion, these results characterize aspects of normal physiology and age-related changes in zebrafish. Furthermore, they suggest that the mechanism through which zebrafish promotes a heat stress response is different from mammals. Taken together, these data support the role for mature zebrafish in comparative studies of aging.